Induction of the marsupial sperm acrosome reaction in vitro by treatment with diacylglycerols

Abstract

The diacylglycerol DiC8 (1,2-dioctanoyl-sn-glycerol; 100 mumol l-1) was found to induce acrosomal loss in 70% of wallaby spermatozoa and 40% of possum spermatozoa after incubation for 120 min. If 10 mumol calcium ionophore l-1 was present, the time required to reach these end points was reduced to 30 and 60 min, respectively. The diacylglycerol OAG (1-oleoyl-2-acetyl-sn-glycerol; 100 mumol l-1) produced some acrosomal loss, particularly when 10 mumol calcium ionophore l-1 was present, but when the ionophore was absent, results were equivocal. The other diacylglycerol tested DOG (1,2-dioleoyl-sn-glycerol; 100 mumol l-1) did not induce significant acrosomal loss in possum or wallaby spermatozoa. The concentration of DiC8 required to induce acrosomal loss in marsupial spermatozoa (50-100 mumol l-1) was relatively high compared with that for placental mammals, suggesting a less specific mode of action than enzyme activation. Of the diacylglycerols tested alone, only DiC8 led to significant loss of motility. In wallabies, this was detectable after incubation for 60 min and in possums after incubation for 120 min. The ultrastructure of the DiC8-induced acrosomal loss was essentially identical to the acrosome reaction described for a broad range of placental mammals. A membrane vesicle shroud covered the acrosomal surface of the sperm head. The vesicles appeared to be formed by multiple point fusions between the plasma membrane and the outer acrosomal membrane. The mechanism of DiC8-induced acrosomal loss remains to be established.(ABSTRACT TRUNCATED AT 250 WORDS)

Diacylglycerol stimulates acrosomal exocytosis by feeding into a PKC- and PLD1-dependent positive loop that continuously supplies phosphatidylinositol 4,5-bisphosphate

“…Addition of DAG or membrane-permeant phorbol esters triggers acrosomal exocytosis in spermatozoa of different species [18][19][20][21]. It has also been demonstrated that DAG formation during acrosomal exocytosis is differentially regulated by transducing pathways activated by physiological agonists [22].…”

Characterisation of fibrous sheath and midpiece fibre network polypeptides of marsupial spermatozoa with a monoclonal antibody

“…Possum and wallaby semen was collected by electroejaculation (Rodger and White, 1978;Rodger and Pollitt, 1981;Sistina et al, 1993). Possum and wallaby spermatozoa were 'swum-up' into phosphate-buffered saline (PBS) as described previously (Mate and Rodger, 1991 [possum]; Sistina et al, 1993 [wallaby]).…”

“…Acrosomal proteins were released from possum spermatozoa, collected from the tail of the epididymis, by treatment with 1,2-dioctanoyl-sn glycerol (DiC 8 , Sigma) as described previously (Sistina et al, 1993). Briefly, possum spermatozoa (10 7 /ml) were treated with 100 µM DiC 8 for 30 min at 37°C.…”

Induction of Thumbtack Sperm During Coculture with Oviduct Epithelial Cell Monolayers in a Marsupial, the Brushtail Possum (Trichosurus vulpecula)1

“…Aliquots (50 l) of unattached sperm were drawn after vigorous pipetting 5 times from the same well at 0, 2, 4, and 6 h of incubation each time and were assessed for sperm concentration, motility, viability, acrosome status, and sperm head-to-tail orientation by standard procedures [25]. Briefly, the acrosome status of the unattached spermatozoa was evaluated after fixation in paraformaldehyde and staining with Bryan's stain, an equal combination of 0.1% (w:v) each of fast green FCF, eosin Y, and flavianic acid (all Sigma Chemical Company) in 1% acetic acid.…”